a. Field of the Invention
The present invention relates to an apparatus for automatically cultivating tissues or cells of a living body, and more particularly to such apparatus capable of performing automatically and successively necessary works for subculture of such tissues or cells under predetermined controlled conditions.
B. Description of the Prior Art
In the present instance, techniques for the culture of such tissues, especially cells of a living body have been of fundamental importance in many fields such as medicine, biology and related fields. Due to the technical difficulty of subculture of the tissues of cells of the living body, especially of a human body, stable cultivated "mass" of such tissues or cells could not be obtained until the technique for cultivating such tissues or cells in a gas-sealed incubator was developed. The tissue or cell culture technique in the gas-sealed incubator is a technique to cultivate tissues or cells of a living body in a predetermined gaseous atmosphere. Thanks to this culture technique, it has become possible to cultivate even the distinctive cells of the human body such as liver cells, nervous cells and hypophysis cells.
Conventional subculture of the tissues or cells of the living body has been performed in the following manner. The cells are diluted first by a nutrient solution so that they float in the nutrient solution. A predetermined number of the cells, together with the nutrient solution, are then put into a container such as a Petri dish. The container containing the predetermined number of the cells and the nutrient solution is disposed in an incubator, the inside of which is maintained in a predetermined gaseous atmosphere. In order to check the multiplication state of the cells in the container, the container is take out from within the incubator after predetermined periods of time have passed, and the multiplication state of the cells is inspected by using a microscope. If it is confirmed by such inspection or observation that the cells in the container have multiplied sufficiently enough so that the container has been filled with the multiplied cells, the container is moved to a clean bench in order to divide the multiplied cells into pieces for further subculture. From within the container in the clean bench the nutrient solution in the container is sucked out of the container by means of a pipet. The multiplied cells left in the container are then rinsed by a buffer solution which is injected in the container and the buffer solution is pipetted out of the container after such rinse. After the cleaning of the cells, an enzyme solution such as trypsin is injected into the container in order to make the multiplied cells that are adhered to the inner bottom surface of the container come off and to divide the cells into pieces. The cells are then separated from the enzyme solution by a centrifugal machine and collected. That is, the cells and enzyme solution in the container are transfussed into a tube of the centrifugal machine so as to be separated from each other, and then the enzyme solution in the tube which exists above the cells is sucked out of the tube. It may not be necessary for some kinds of cells to use the centrifugal machine for the purpose of separating the cells and the enzyme solution from each other. In this case, the enzyme solution is sucked out of the container immediately before the adhered cells are liberated by the enzyme solution and the cells are subjected to the next process.
After the enzyme solution has been sucked out of the tube of the centrifugal machine, a fresh nutrient solution is charged into the tube so that the cells in the tube are diluted in the predetermined density of dilution, and this nutrient solution containing the cells divided into pieces float is distributed into empty containers of a predetermined quantity. The container which contains a predetermined number of cells and nutrient solution is then disposed in the incubator, the inside of which is maintained at a predetermined atmosphere, so that the cells in the container are again cultivated.
However, the conventional cultivating manner described hereinbefore has the following disadvantages. One of such disadvantages is that the cultivating container must be often taken out from within the incubator into the air in order to check the multiplication state of the tissues or cells by using the microscope. Therefore, the environment for the culture i.e. gaseous atmosphere, temperature and humidity is greatly changed every time the container is taken out from within the incubator into the air, and this environmental change often causes subtle variations or changes in the tissues or cells in the container. In addition to this drawback, the tissues or cells in the container are contaminated by bacteria or germs contained in the air.
Furthermore, since the necessary operations for subculture, i.e. separating the cells from the enzyme solution, collecting the cells, diluting the cells by the nutrient solution, charging the cells and nutrient solution into the empty container and etc., are munually performed within the clean bench by a skilled technician in accordance with the results of the observation by the microscope, the property of characteristics of the cultivated cells are directly affected by the technician's ability. That is, the property or characteristics of the cultivated tissues or cells depend on the experience and skill of the technician.
Therefore, it is impossible by the conventional cultivating manner to obtain a unified or standardized cultivated tissues or cells.
As the result of the fact that the standardization or unification of the cell culture is very difficult for the reasons described above, opposite conclusions are sometimes obtained by different scientists who have respectively studied the same theme. Thus, the scientist must spare his considerable energies for the cell culture which is not his essential work, instead of devoting his energies to his original medical or biological studies. This is a common worry among the scientists of various fields, and thus the unification or standardization of the tissue or cell culture has been highly desired